Anti-Inflienza drug screening targetted by cap-snatching mechanism

• Project/Area Number: 15K08502
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Virology
• Research Institution: Kitasato University
• Principal Investigator: SHIBAGAKI Yoshio 北里大学, 薬学部, 講師 (90235565)
• Project Period (FY): 2015-04-01 – 2018-03-31
• Project Status: Completed (Fiscal Year 2017)
• Budget Amount: ¥4,940,000 (Direct Cost: ¥3,800,000、Indirect Cost: ¥1,140,000); Fiscal Year 2017: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2016: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000); Fiscal Year 2015: ¥1,820,000 (Direct Cost: ¥1,400,000、Indirect Cost: ¥420,000)
• Keywords: cap structure / antiinfluenza drug / cap-snatching / キャップスナッチング / 抗インフルエンザ薬 / リード化合物スクリーニング / キャップ構造 / インフルエンザ転写反応 / 微生物代謝物スクリーニング / Cap-snatching阻害活性 / 微生物ライブラリースクリーニング

Outline of Final Research Achievements

This cap-dependent endonuclease activity termed as cap snatching may provide a unique target for novel anti-influenza viral agents. For the screening of candidate inhibitors for cap-snatching activity, it is essential to establish a method to efficiently produce Cap1-RNA substrate and a convenient assay system for cap-snatching activity. A short 3’-biotinylated RNA oligonucleotide was prepared by an in vitro transcription system utilizing T7 RNA polymerase. Purified 3’-biotinylated RNA oligonucleotide by C18 cartridge column was subjected to a sequential capping reaction with recombinant enzymes of vaccinia virus D1R containing with RNA 5’-triphosphatase and mRNA guanylyltransferase, yeast
ABD1 as (guanine-N7)-methyltransferase, and VP39 as (nucleoside-2'-O-)-methyltransferase. Cap-snatching activity could be analyzed a pull-down assay based on the streptavidin-biotin interaction, yielding results consistent with those obtained by polyacrylamide gel electrophoresis.

Research Products

• [Journal Article] Phosphoproteomic Analysis Identifies Signaling Pathways Regulated by Curcumin in Human Colon Cancer Cells (2017)
  • Author(s): TATSUHIRO SATO, YUTAKA HIGUCHI, YOSHIO SHIBAGAKI and SEISUKE HATTORI
  • Journal Title: Anticancer Research Volume: 37 Issue: 9 Pages: 4789-4798
  • DOI: 10.21873/anticanres.11885
  • Peer Reviewed / Open Access
• [Journal Article] Mechanistic insights into ectodomain shedding: susceptibility of CADM1 adhesion molecule is determined by alternative splicing and O-glycosylation (2017)
  • Author(s): Kyoko Shirakabe, Takuya Omura, Yoshio Shibagaki, Emiko Mihara, Keiichi Homma, Yukinari Kato, Akihiko Yoshimura, Yoshinori Murakami, Junichi Takagi, Seisuke Hattori & Yoshihiro Ogawa
  • Journal Title: Scientific reports Volume: 7 Issue: 1 Pages: 46174-46174
  • DOI: 10.1038/srep46174
  • Peer Reviewed / Open Access
• [Journal Article] Src Family Tyrosine Kinase Signaling Regulates FilGAP through Association with RBM10 (2016)
  • Author(s): Hazuki Yamada, Koji Tsutsumi, Yuki Nakazawa, Yoshio Shibagaki, Seisuke Hattori, Yasutaka Ohta
  • Journal Title: PloS one Volume: 11 Issue: 1 Pages: e0146593-e0146593
  • DOI: 10.1371/journal.pone.0146593
  • Peer Reviewed / Open Access
• [Journal Article] CCT2 https://www-kofu.jsps.go.jp/kofu1/images/bt_add_small.gifMutations Evoke Leber Congenital Amaurosis due to Chaperone Complex Instability. (2016)
  • Author(s): Minegishi Y, Sheng X, Yoshitake K, Sergeev Y, Iejima D, Shibagaki Y, Monma N, Ikeo K, Furuno M, Zhuang W, Liu Y, Rong W, Hattori A, Iwata T.
  • Journal Title: Sci Rep Volume: 6 Issue: 1 Pages: 33742-33742
  • DOI: 10.1038/srep33742
  • Peer Reviewed / Open Access
• [Journal Article] Mammalian target of rapamycin (mTOR) complex 2 regulates filamin A-dependent focal adhesion dynamics and cell migration (2016)
  • Author(s): Tatsuhiro Sato, Junko Ishii, Yuki Ota, Eri Sasaki, Yoshio Shibagaki, Seisuke Hattor
  • Journal Title: Genes to Cells Volume: 未定 Issue: 6 Pages: 579-593
  • DOI: 10.1111/gtc.12366
  • Peer Reviewed / Open Access / Acknowledgement Compliant
• [Journal Article] Identification of 4-Trimethylaminobutyraldehyde Dehydrogenase (TMABA-DH) as a Candidate Serum Autoantibody Target for Kawasaki Disease (2015)
  • Author(s): Atsuko Matsunaga, Yutaka Harita, Yoshio Shibagaki, Nobutaka Shimizu, Kazuhiko Shibuya, Hiroshi Ono, Hitoshi Kato, Takashi Sekine, Naoko Sakamoto, Takashi Igarashi, Seisuke Hattori
  • Journal Title: PLoS One Volume: 10 Issue: 5 Pages: e0128189-e0128189
  • DOI: 10.1371/journal.pone.0128189
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Rheb protein binds CAD (carbamoyl-phosphate synthetase 2, aspartate transcarbamoylase, and dihydroorotase) protein in a GTP- and effector domain-dependent manner and influences its cellular localization and carbamoyl-phosphate synthetase (CPSase) activity. (2015)
  • Author(s): Sato T, Akasu H, Shimono W, Matsu C, Fujiwara Y, Shibagaki Y, Heard JJ, Tamanoi F, Hattori S.
  • Journal Title: J Biol Chem. Volume: 290 Issue: 2 Pages: 1096-1105
  • DOI: 10.1074/jbc.m114.592402
  • Peer Reviewed / Open Access
• [Presentation] Cap-snatching をターゲットとした抗インフルエンザ薬スクリーニング (2016)
  • Author(s): 柴垣芳夫
  • Organizer: 第39日本分子生物学会年会
  • Place of Presentation: 横浜
  • Year and Date: 2016-11-30
• [Presentation] Cap-snatching 反応をターゲットとした抗インフルエンザ薬スクリーニング (2015)
  • Author(s): 柴垣芳夫1)、浅野 恵美子1)、兒玉 朋子1)、野村 大成1)、内田 龍児2)、供田 洋2)、服部 成介
  • Organizer: 日本分子生物学会
  • Place of Presentation: 神戸ポートアイランド (兵庫県神戸市中央区)
  • Year and Date: 2015-12-01